Effect Of ?-Asarone To Protect PC12 Cells Was Injuried By Glu And The Expression Of NMDAR Subnits

ObjectiveTo discuss the protective effects of ?-asarone for PC12 cells injuried by glutamate and the expression of NMDA receptor subnits and synaptophysin.MethodsPC12 cells divided into normal group,model group and treat group.The normal group was cultured with serum-free DMEM medium;The model group was cultured with serum-free DMEM containing 50mM glutamic acid;the treatment group was cultured with 10 ? M of asarone and 50mM containing glutamate in serum-free DMEM medium.To observe the cells morphology under electron microsc-ope and CCK-8 to test its vitality after 2h,and immunohistochemistry to detect the expression of synaptophysin(SYP).Flow cytometry to detect the change of intracellular Ca2+,mitochondrial membrane potential and apoptosis;Immunofluorescence technique to detect the SYP expression in each cell group;PCR technique to detect the expression of NMDA receptor subtypes gene in different group of cells.ResultsTo compare with model group,the cell vigor was increased(P<0.01),to compare with the treatment group,the cell vigor of the treatment group was obviously higher than the model group(P<0.01),Compared with the treatment group,the cell vigor of the normal group was higher than the treatment group(P<0.01).Model group Intracellular Ca2+ concentration was increased to compare with normal group(P<0.01);while lowed with treat group(P<0.05);Compared with the normal group,the Ca2+ concentration in the treatment group was not statistically significant(P>0.05).Model group mitochondrial transmembrane potential was lower than normal group(P<0.01),treatment group mitochondrial transmembrane potential was higher than model group(P<0.05);To compare with the normal group,the mitochondrial membrane potential of the normal group was higher than that of the treatment group(P<0.01).Model group apopotosis rate was higher than normal group(P<0.01):while treat group apopotosis rate was lower than model group(P<0.05);To compare with the normal group,the normal group of apoptosis rate was lower than that of the treatment group(P<0.01).Compared with the normal group,the SYP fluorescence intensity in the model group was weaker than the normal group(P<0.01),and the fluorescence intensity of SYP in the treatment group was stronger than the model group(P<0.05),the number of SYP positive cells in normal group to compare with the model group was increased(P<0.01),it was decreased the number of SYP positive cells in model group to compare with the treatment group(P<0.05).Compared with the normal group,there was no significant difference in the number of SYP positive cells in the treatment group(P>0.05).Compared with the normal group,the expression of NR1 mRNA in the model group were signifi-cantly lower than the normal group(P<0.01),and the expression of NR2A mRNA in model group was obviously lower than in normal group(P<0.01);compared with the model group,the expression of NR1 mRNA cells in treatment group was significantly higher than that in the model group(P<0.01),the expression of NR2A in the treatment group was high to compare with the model group(P<0.05).Compared with the normal group,the expression of NR2B mRNA in the model group higher than in normal group(P<0.05),the expression of NR2C mRNA in the model group was significantly lower than normal group(P<0.01);To compare with the model group,NR2B expression of mRNA in the treatment was higher than the model group(P<0.05),the expression of NR2C in treatment group and model of high group(P<0.05).Compared with the normal group,the expression of NR2D mRNA in the model groupis no statistical significance(P>0.05),while the expression of NR3A in the model group was significantly lower than normal group(f<0.01);the NR2D mRNA expression in the treatment group were not stati-stically significant to compare with the model group(P>0.05),the expression of NR3A in treatment group was high in the model group(P<0.05).To compare with the normal group,the expression of NR1,NR2B,NR2C,NR2D,NR3A mRNA in the treatment group was not statistically significant(P>0.05),while the expression of NR2A mRNA was lower than that of the normal group(P<0.05).However,we did not detect the expression of NR3B mRNA in each group of cells.Conclusion?-asarone had a protective effect on the PC12 cells injuried by GLU,it could increase the SYP expression and regulate the NMDAR subnits to express,but we still unknown the mechanism and need further to study.